Intraocular Pressure and Medication Burden With Cataract Surgery Alone, or Cataract Surgery Combined With Trabecular Bypass or Goniotomy.

PRCIS: When compared with cataract surgery in glaucoma patients, trabecular micro-bypass and goniotomy resulted in a large decrease in the incidence of intraocular pressure (IOP) spikes, a modest effect on IOP, and a minimal effect on medication burden. PURPOSE: To compare changes in IOP and ocular hypotensive medications in 3 surgical cohorts: cataract surgery, cataract surgery with trabecular micro-bypass (cataract/trabecular), and cataract surgery with goniotomy (cataract/goniotomy). MATERIALS AND METHODS: We included 138 eyes diagnosed with open-angle glaucoma: (1) 84 eyes with cataract surgery alone, (2) 25 eyes with cataract/trabecular surgery, and (3) 29 eyes with cataract/goniotomy surgery. We compared the groups for postoperative IOP and the number of ocular hypotensive medications. We adjusted for preoperative IOP, and preoperative and postoperative number of ocular hypotensive medications. We defined an IOP spike as IOP ≥21 mm Hg and 10 mm Hg higher than preoperative on postoperative day 1. RESULTS: All 3 surgeries showed a decrease in IOP (P≤0.004) and medication burden (P≤0.001) at 3 and 6 months postoperatively when compared with their own preoperative baselines. When compared with cataract surgery alone, cataract/trabecular and cataract/goniotomy had similar IOP lowering at 1 month postoperatively, and variable results at 3 and 6 months. The change in ocular hypotensive medications was not statistically different between the surgical groups at any postoperative visit. Cataract/trabecular and cataract/goniotomy decreased IOP on postoperative day 1, and had relative risk reduction of ~70% for IOP spikes (P≤0.001 for both). CONCLUSION: Trabecular micro-bypass and goniotomy when added to cataract surgery resulted in a large decrease in IOP spikes, a modest effect on IOP, and a minimal effect on medication burden when compared with cataract surgery alone in glaucoma patients.

Predictive Value of Excellent Uncorrected Visual Acuity Post-Operative Day One After Cataract Surgery.

PURPOSE: Patients and physicians are often pleased when uncorrected visual acuity (UCVA) on post-operative day 1 (POD1) after cataract surgery is 20/20. Unfortunately, this UCVA does not always last. This article aims to investigate the relationship between excellent uncorrected visual acuity on post-operative day 1 and final post-operative UCVA after uncomplicated cataract surgery. PATIENTS AND METHODS: The medical records of patients who had undergone uncomplicated cataract surgery between 2012 and 2017 were assessed. UCVA on POD1 and final UCVA were obtained for patients who had a final best-corrected visual acuity of 20/20 or better. RESULTS: Of 309 patients with UCVA of 20/20 on POD 1, 62.4% maintained 20/20 and 87.4% maintained 20/25 or better as their final uncorrected visual outcome. Of 204 patients with UCVA of 20/25 on POD 1, 44.1% achieved 20/20 and 69.6% maintained 20/25 or better as their final uncorrected visual outcome. Patients with 20/20 UCVA on POD1 were more likely to have a better final UCVA compared with those who were 20/25 on POD1. Of the 531 patients with UCVA of 20/25 or better on POD1, 20% had final UCVA worse than 20/25 with 4% losing more than 2 lines for their final UCVA. CONCLUSION: The majority of patients with 20/20 UCVA on POD1 after cataract surgery maintained excellent UCVA as their final visual outcome. However, a significant percentage of these patients experienced a decrease in UCVA over the course of the postoperative period.

Brinzolamide-induced Follicular Conjunctivitis.

We report a case of a patient who developed a severe case of follicular conjunctivitis from brinzolamide after 1.5 years of consistent use. This patient was rechallenged again after resolution of the follicles and subsequently redeveloped similar conjunctivitis. This is the first confirmed reported case of follicular conjunctivitis from brinzolamide use.

Transmission of Donor-Derived Breast Carcinoma as a Recurrent Mass in a Keratolimbal Allograft.

PURPOSE: To report a case of local transmission of invasive lobular carcinoma from a donor to a recipient in a keratolimbal allograft after cessation of systemic immunosuppressive therapy. METHODS: This is a case report including the clinicopathologic findings. Sections of the donor breast tumor and recipient conjunctival lesions were stained with hematoxylin and eosin. Immunohistochemical studies were performed using pancytokeratin, CK7, CK20, CAM 5.2, CD138, TTF1, estrogen receptor, progesterone receptor, GATA-3, GCDFP-15, and mammaglobin. Polymerase chain reaction-based DNA profiling of tumor cells was performed. RESULTS: Histopathologic examination revealed an infiltrate of atypical cells with large hyperchromatic nuclei consistent with carcinoma. Immunohistochemical analysis showed pancytokeratin, CK7, CAM 5.2, GATA-3, and estrogen receptor positivity and progesterone receptor absence, consistent with the previously determined phenotype of the donor's breast carcinoma. Results of polymerase chain reaction analysis were also consistent with the donor's tumor. After reduced dosing of tacrolimus and mycophenolate mofetil, 2 limbal tumors occurred in the recipient. The immunosuppressive treatment had been stopped completely before the appearance of the third lesion. The recipient had no history of malignancy, and she had routine screenings for breast cancer. CONCLUSIONS: We report a case of donor-derived breast carcinoma in a keratolimbal allograft recipient. The grafted tissue harbored donor-derived tumor cells for more than 4 years after surgery even after systemic immunosuppression was discontinued. Although no similar reports of tumor transfer could be found in the literature, this case suggests the need for increased stringency in donor selection and heightened surveillance for such tumor transmission.

Hereditary Benign Intraepithelial Dyskeratosis: Report of a Case and Re-examination of the Evidence for Locus Heterogeneity.

BACKGROUND: Hereditary benign intraepithelial dyskeratosis (HBID) is a rare autosomal-dominant disorder of the conjunctiva and oral mucosa first described in and predominantly affecting descendents of Haliwa-Saponi Native Americans. We report a spontaneous case of histopathologically-confirmed HBID affecting an individual not of Native American ancestry. MATERIALS AND METHODS: Report of a case with histopathologic examination of an excised conjunctival specimen as well as molecular and cytogenetic analysis. RESULTS: A Caucasian boy with a history of oral lesions and conjunctival injection from birth developed bilateral corneal opacities at age 5 and underwent penetrating keratoplasty, with recurrence of the corneal opacification shortly after surgery. Examination of a conjunctival biopsy specimen revealed features consistent with HBID. Copy number variant (CNV) analysis revealed a de novo 4q35 duplication that overlapped the duplication previously associated with HBID, although no genes were identified in the common interval. NLRP1 gene sequencing failed to reveal a presumed pathogenic variant. CONCLUSIONS: HBID may develop de novo in individuals who are not of Native American ancestry. The absence of coding regions in a duplicated region of 4q35 common to both the individual that we report and previously associated with HBID raises questions regarding the significance of this CNV in the pathogenesis of HBID.

X-linked Megalocornea Associated with the Novel CHRDL1 Gene Mutation p.(Pro56Leu*8).

BACKGROUND: The genetic basis of X-linked megalocornea (MGC1) was reported in 2012 to be caused by mutations in the CHRDL1 gene. We sought to confirm that mutations in CHRDL1 are associated with MGC1 in a previously unreported pedigree. MATERIALS AND METHODS: Slit lamp examination, corneal pachymetry, corneal topography and DNA collection for screening of the CHRDL1 gene were performed for members of an affected family. RESULTS: Examination of a woman and her four sons, ranging in age between 3 and 15 years, demonstrated horizontal corneal diameters of 14 mm in three of the four sons and a normal corneal diameter of 12 mm in the mother and other son. Central corneal thickness in the individuals with enlarged corneal diameters averaged 474 microns, compared to 604 microns in their unaffected brother. Corneal topographic imaging demonstrated an average K value of 44.4 D in the affected individuals compared with 41.6 D in their unaffected sibling. Screening of the CHRDL1 gene demonstrated the novel hemizygous frameshift mutation c.167delC (p.(Pro56Leu*8)) in exon 3 in the affected individuals and in the heterozygous state in their mother. This mutation was not present in the unaffected brother or in unrelated controls. CONCLUSION: We provide the initial confirmation that X-linked megalocornea is associated with mutations in the CHRDL1 gene.

Separation of porcine parvovirus from bovine serum albumin using PEG-salt aqueous two-phase system.

Vaccine production faces a challenge in adopting conventional downstream processing steps that can efficiently purify large viral particles. Some major issues that plague vaccine purification are purity, potency, and quality. The industry currently considers 30% as an acceptable virus recovery for a vaccine purification process, including all downstream processes, whereas antibody recovery from CHO cell culture is generally around 80-85%. A platform technology with an improved virus recovery would revolutionize vaccine production. In a quest to fulfill this goal, we have been exploring aqueous two-phase systems (ATPSs) as an optional mechanism to purify virus. ATPS has been unable to gain wide implementation mainly due to loss of virus infectivity, co-purification of proteins, and difficulty of polymer recycling. Non-enveloped viruses are chemically resistant enough to withstand the high polymer and salt concentrations that are required for effective ATPS separations. We used infectious porcine parvovirus (PPV), a non-enveloped, DNA virus as a model virus to test and develop an ATPS separation method. We successfully tackled two of the three main disadvantages of ATPS previously stated; we achieved a high infectious yield of 64% in a PEG-citrate ATPS process while separating out the main contaminate protein, bovine serum albumin (BSA). The most dominant forces in the separation were biomolecule charge, virus surface hydrophobicity, and the ATPS surface tension. Highly hydrophobic viruses are likely to benefit from the discovered ATPS for high-purity vaccine production and ease of implementation.

Rate of environmental change determines stress response specificity.

Cells use general stress response pathways to activate diverse target genes in response to a variety of stresses. However, general stress responses coexist with more specific pathways that are activated by individual stresses, provoking the fundamental question of whether and how cells control the generality or specificity of their response to a particular stress. Here we address this issue using quantitative time-lapse microscopy of the Bacillus subtilis environmental stress response, mediated by σ(B). We analyzed σ(B) activation in response to stresses such as salt and ethanol imposed at varying rates of increase. Dynamically, σ(B) responded to these stresses with a single adaptive activity pulse, whose amplitude depended on the rate at which the stress increased. This rate-responsive behavior can be understood from mathematical modeling of a key negative feedback loop in the underlying regulatory circuit. Using RNAseq we analyzed the effects of both rapid and gradual increases of ethanol and salt stress across the genome. Because of the rate responsiveness of σ(B) activation, salt and ethanol regulons overlap under rapid, but not gradual, increases in stress. Thus, the cell responds specifically to individual stresses that appear gradually, while using σ(B) to broaden the cellular response under more rapidly deteriorating conditions. Such dynamic control of specificity could be a critical function of other general stress response pathways.

Measuring single-cell gene expression dynamics in bacteria using fluorescence time-lapse microscopy.

Quantitative single-cell time-lapse microscopy is a powerful method for analyzing gene circuit dynamics and heterogeneous cell behavior. We describe the application of this method to imaging bacteria by using an automated microscopy system. This protocol has been used to analyze sporulation and competence differentiation in Bacillus subtilis, and to quantify gene regulation and its fluctuations in individual Escherichia coli cells. The protocol involves seeding and growing bacteria on small agarose pads and imaging the resulting microcolonies. Images are then reviewed and analyzed using our laboratory's custom MATLAB analysis code, which segments and tracks cells in a frame-to-frame method. This process yields quantitative expression data on cell lineages, which can illustrate dynamic expression profiles and facilitate mathematical models of gene circuits. With fast-growing bacteria, such as E. coli or B. subtilis, image acquisition can be completed in 1 d, with an additional 1-2 d for progressing through the analysis procedure.

Stochastic pulse regulation in bacterial stress response.

Gene regulatory circuits can use dynamic, and even stochastic, strategies to respond to environmental conditions. We examined activation of the general stress response mediated by the alternative sigma factor, σ(B), in individual Bacillus subtilis cells. We observed that energy stress activates σ(B) in discrete stochastic pulses, with increasing levels of stress leading to higher pulse frequencies. By perturbing and rewiring the endogenous system, we found that this behavior results from three key features of the σ(B) circuit: an ultrasensitive phosphorylation switch; stochasticity ("noise"), which activates that switch; and a mixed (positive and negative) transcriptional feedback, which can both amplify a pulse and switch it off. Together, these results show how prokaryotes encode signals using stochastic pulse frequency modulation through a compact regulatory architecture.

Mixed messages: how bacteria resolve conflicting signals.

An elegant new study by Bollenbach and Kishony (2011) in this issue of Molecular Cell shows how bacteria resolve the apparent conflicts created when they face two signals with opposite effects on gene expression.

Accurate prediction of gene feedback circuit behavior from component properties.

A basic assumption underlying synthetic biology is that analysis of genetic circuit elements, such as regulatory proteins and promoters, can be used to understand and predict the behavior of circuits containing those elements. To test this assumption, we used time-lapse fluorescence microscopy to quantitatively analyze two autoregulatory negative feedback circuits. By measuring the gene regulation functions of the corresponding repressor-promoter interactions, we accurately predicted the expression level of the autoregulatory feedback loops, in molecular units. This demonstration that quantitative characterization of regulatory elements can predict the behavior of genetic circuits supports a fundamental requirement of synthetic biology.

Gene regulation at the single-cell level.

The quantitative relation between transcription factor concentrations and the rate of protein production from downstream genes is central to the function of genetic networks. Here we show that this relation, which we call the gene regulation function (GRF), fluctuates dynamically in individual living cells, thereby limiting the accuracy with which transcriptional genetic circuits can transfer signals. Using fluorescent reporter genes and fusion proteins, we characterized the bacteriophage lambda promoter P(R) in Escherichia coli. A novel technique based on binomial errors in protein partitioning enabled calibration of in vivo biochemical parameters in molecular units. We found that protein production rates fluctuate over a time scale of about one cell cycle, while intrinsic noise decays rapidly. Thus, biochemical parameters, noise, and slowly varying cellular states together determine the effective single-cell GRF. These results can form a basis for quantitative modeling of natural gene circuits and for design of synthetic ones.